O2 sensing is preserved in mice lacking the gp91 phox subunit of NADPH oxidase

O₂ sensing is preserved in mice lacking the gp91 phox subunit of NADPH oxidase

^*Department of Medicine, Division of Cardiology, and ^§Department of Pathology, University of Alberta, Edmonton, Canada T69 2B7; ^†Department of Medicine, Veterans Administration Medical Center, Minneapolis, MN 55417; ^‡Department of Physiology, University of Minnesota, Minneapolis, MN 55455; and ^¶Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN 46202

Communicated by Paul B. Beeson, University of Washington, Redmond, WA (received for review January 15, 1999)

Abstract

The rapid response to hypoxia in the pulmonary artery (PA), carotid body, and ductus arteriosus is partially mediated by O₂-responsive K⁺ channels. K⁺ channels in PA smooth muscle cells (SMCs) are inhibited by hypoxia, causing membrane depolarization, increased cytosolic calcium, and hypoxic pulmonary vasoconstriction. We hypothesize that the K⁺ channels are not themselves “O₂ sensors” but rather respond to the reduced redox state created by hypoxic inhibition of candidate O₂ sensors (NADPH oxidase or the mitochondrial electron transport chain). Both pathways shuttle electrons from donors, down a redox gradient, to O₂. Hypoxia inhibits these pathways, decreasing radical production and causing cytosolic accumulation of unused, reduced, freely diffusible electron donors. PASMC K⁺ channels are redox responsive, opening when oxidized and closing when reduced. Inhibitors of NADPH oxidase (diphenyleneiodonium) and mitochondrial complex 1 (rotenone) both inhibit PASMC whole-cell K⁺ current but lack the specificity to identify the O₂-sensor pathway. We used mice lacking the gp91 subunit of NADPH oxidase [chronic granulomatous disease (CGD) mice] to assess the hypothesis that NADPH oxidase is a PA O₂-sensor. In wild-type lungs, gp91 phox and p22 phox subunits are present (relative expression: macrophages > airways and veins > PASMCs). Deletion of gp91 phox did not alter p22 phox expression but severely inhibited activated O₂ species production. Nonetheless, hypoxia caused identical inhibition of whole-cell K⁺ current (in PASMCs) and hypoxic pulmonary vasoconstriction (in isolated lungs) from CGD vs. wild-type mice. Rotenone vasoconstriction was preserved in CGD mice, consistent with a role for the mitochondrial electron transport chain in O₂ sensing. NADPH oxidase, though a major source of lung radical production, is not the pulmonary vascular O₂ sensor in mice.

Footnotes

↵ ‖ To whom reprint requests should be addressed at: Veterans Administration Medical Center (111C), One Veterans Drive, Minneapolis, MN 55417. e-mail: weirx002{at}maroon.tc.umn.edu.
ABBREVIATIONS:

PA,

pulmonary artery;

SMC,

smooth muscle cell;

CGD,

chronic granulomatous disease;

ETC,

electron transport chain;

HPV,

hypoxic pulmonary vasoconstriction;

Kv,

voltage-gated potassium channel;

IK, whole-cell K+ current,

EM, membrane potential;

GSH/GSSG reduced/oxidized glutathione,

IBTX, iberiotoxin;

AOS,

activated O2 species;

PMA,

phorbol myristate acetate;

DPI,

diphenyleneiodonium;

PVR,

pulmonary vascular resistance;

PAP,

pulmonary artery pressure;

AII,

angiotensin II;

4-AP,

4-aminopyridine